Project description:Deficiency of deacetylase SIRT1 or its downstream target PRRX1 promotes breast cancer stemness and metastasis. To explore the mechnisms of SIRT1 and its downstream target PRRX1 in breast cancer, we analysed the gene expressions in breast cancer BT549 cells with SIRT1 knockout or PRRX1 knockout or not.
Project description:NAD+ levels decline with age and in certain disease conditions. NAD+ precursors have been shown to stimulate NAD+ biosynthesis and ameliorate various age-associated diseases in mouse models. However, NAD+ metabolism is complicated in cancer and its role in triple-negative breast cancer (TNBC) remains elusive. Here, we show that NAD+ supplement suppresses tumor metastasis in a TNBC orthotopic patient-derived xenograft (PDX) model. Sirtuin1 lysine deacetylase (SIRT1) is required for the effects since SIRT1 knockdown blocks NAD+-suppressed tumor metastasis. Overexpression of SIRT1 effectively impairs the metastatic potential of TNBC. Importantly, the interaction between SIRT1 and p66Shc causes the deacetylation and functional inactivation of p66Shc, which inhibits epithelial-mesenchymal transition (EMT). Overall, we demonstrate that NAD+ supplementation executes its anti-tumor function via activating the SIRT1-p66Shc axis, which highlights the preventive and therapeutic potential of SIRT1 activators as effective interventions for TNBC.
Project description:SIRT1 is a nuclear NAD+-dependent protein deacetylase. Expression microarray analysis was used to study the effect of SIRT1 knockdown on gene expression in MCF-7 breast cancer cells. Experiment Overall Design: SIRT1 stable knockdown was achieved using two retroviral shRNA constructs. An shRNA directed against Luciferase was used to generate the Luc control cells. Three independent biological replicates with matching Luc controls were analyzed using Affymetrix U133A 2.0 microarrays. Experiment Overall Design: These 6 samples were part of a 15-sample microarray analysis (GSE12971) examining expression regulation by SIRT1, PARP-1, PARG and macroH2A. All 15 samples were included for the following data normalization steps: data sets obtained using the GCOS software were grouped based on date of experiment and adjusted for batch effect using the parametric empirical Bayes method (Combat R); all values < 0.01 were adjusted to 0.01; the data were log2 transformed, median centered for each array, and median centered for each gene.
Project description:Analysis of BGC-823 gastric cancer cells with SIRT1 overexpression or knockdown. SIRT1, a NAD+-dependent protein deacetylase, exerts inhibitory effects on migration and invasion of gastric cancer. Results provide insight into the role of SIRT1 in the metastasis of gastric cancer.
Project description:Affymetrix expression arrays were used to compare expression patterns upon knockdown of PARP-1, PARG, SIRT1, or macroH2A in comparison to Luciferase control. Total RNA isolated from three independently generated control-matched MCF-7 knockdown cell lines targeting Luciferase (control), PARP-1, PARG, SIRT1, and macroH2A was used to probe Affymetrix Human U133A 2.0 Expression Microarrays. The raw array data was processed by Affymetrix GeneChip Operating Software (GCOS) to obtain detection calls (ABS_CALL) and signal values (VALUE_2). The signals were then normalized by scaling to a target value of 500 using GCOS. To adjust for batch effects due to day-to-day differences in RNA isolations, the empirical Bayes method was applied to the data set. After adjusting any values less than 0.01 to 0.01, the data was log2 transformed, median centered for each array, and median centered for each individual probe set (VALUE). Filters were then applied to obtain final gene lists.
Project description:FAT1, a protocadherin, is among the most frequently mutated genes in human cancers1-5. However, the role and the molecular mechanisms by which FAT1 mutations control tumour initiation and progression are poorly understood. In the present study we used different mouse cancer models including skin squamous cell carcinoma (SCC) and lung tumours we found that Fat1 deletion accelerated tumour initiation and malignant progression and promoted hybrid epithelial to mesenchymal transition (EMT) phenotype. This hybrid EMT state was also found in FAT1 mutated human SCCs. Fat1 deleted skin SCCs presented increased tumour stemness and spontaneous metastasis. Transcriptional and chromatin profiling revealed that Yap1 and Sox2 are involved in promoting and stabilizing hybrid EMT state. To unravel the molecular mechanisms by which FAT1 (a protein located on the plasma membrane), lead to the transcriptional changes, we performed phosphor-proteomic analysis of A388 FAT1 WT human SCC cell lines and A388 FAR1 CRISPR KO. This analysis revealed that FAT1 loss of function activates a CAMK2/CD44/SRC axis that promotes YAP/ZEB1 nuclear translocation and stimulates the mesenchymal state, as well as a CAMK2-EZH2 axis that promotes activation of SOX2, which sustains the epithelial state. This comprehensive analysis also identified drug resistance and vulnerabilities in FAT1 deficient tumours with important implications for cancer therapy. Altogether, our studies revealed that Fat1 loss of function promotes tumour initiation, progression, invasiveness, stemness and metastasis through the induction of a hybrid EMT state.
Project description:Pancreatic cancer is a common malignant tumor with poor prognosis. Accumulating evidence indicates that sirtuin 1 (SIRT1) exerts biological functions in various cancers.SIRT1 plays an important role in tumorigenesis, development, and drug resistance by blocking aging and apoptosis, also promoting cell growth and angiogenesis.However, how SIRT1 functions in the development of pancreatic cancer stem cells (CSCs) is currently poorly understood. Here, we showed that SIRT1 is involved in regulating various pathways closely related to tumor growth, autophagy and stemness, as well as promoting pancreatic CSC development.
Project description:Pancreatic cancer is a common malignant tumor with poor prognosis. Accumulating evidence indicates that sirtuin 1 (SIRT1) exerts biological functions in various cancers.SIRT1 plays an important role in tumorigenesis, development, and drug resistance by blocking aging and apoptosis, also promoting cell growth and angiogenesis.However, how SIRT1 functions in the development of pancreatic cancer stem cells (CSCs) is currently poorly understood. Here, we showed that SIRT1 inhibits the differentiation of cancer cells by inhibiting various genes expression , including GRHL3, thus promoting the stemness and tumorigenesis of pancreatic cancer.
Project description:Sirtuin-1 (Sirt1), a class III histone/protein deacetylase is central to cellular metabolism, stress responses and aging, but its contributions to various host immune functions have been little investigated. To study the role of Sirt1 in T-cell functions, we undertook targeted deletions by mating mice with a floxed Sirt1 gene to mice expressing CD4-cre or Foxp3-cre recombinase, respectively. We found that Sirt1 deletion left conventional T-effector cell activation, proliferation and cytokine production largely unaltered. However, Sirt1 targeting promoted the expression and acetylation of Foxp3, a key transcription factor in T-regulatory (Treg) cells, and increased Treg suppressive functions in vitro and in vivo. Consistent with these data, mice with targeted deletions of Sirt1 in either CD4+ T-cells or Foxp3+ Treg cells exhibited prolonged survival of MHC-mismatched cardiac allografts. Allografts in Sirt1 targeted recipients showed long-term preservation of myocardial histology and infiltration by Foxp3+ Treg cells. Comparable results were seen in wild-type allograft recipients treated with Sirt1 inhibitors, such as EX-527 and splitomicin. Hence, Sirt1 may inhibit Treg functions and its targeting may have therapeutic value in autoimmunity and transplantation. RNA from three independent samples from magnetically separated CD4+CD25+ Treg of Sirt1 knock out, compared to wild type (C57BL6) control